Enhancers are critical for orchestrating correct development and differentiation and many diseases are associated with changes in enhancer activity. There is a strong desire in the field to identify the direct target promoters of disease-associated enhancers. The development of a robust method by which to do so would greatly increase our mechanistic understanding of gene regulation and, importantly, would enhance our ability to understand the functional significance of distal regulatory elements linked to increased risk for human diseases. Two approaches to experimentally link enhancers to target promoters have dominated the field over the last several years; these are a) using a genomic nuclease to delete an enhancer followed by RNA-seq to identify genes with altered regulation and b) identifying a target gene of an enhancer using 3D looping detection methods. However, these methods have not been as useful as anticipated. Clearly, what is needed is a new method that enables investigators to identify target promoters of enhancers that does not rely on the existence of a stable enhancer-promoter loop and that is not hindered by the problem of enhancer redundancy. We propose to develop a technology termed ?promoter tagging? which is based on the concept of using ?molecular memory? to identify transient looping between enhancers and promoters; we will compare methods that modify genomic DNA (Aim 1), histone H3 (Aim 2), or chromatin-bound proteins (Aim 3) at both the enhancer region and at a target promoter that has recurrent, transient interactions with the enhancer. Our studies will provide innovative approaches to solve an important problem in the field of genomic regulation. The ability to identify transient looping between enhancers and promoters is of enormous significance because of the substantial evidence that many regulatory factors act through a hit-and-run mechanism. Importantly, our proposed new technology can be used by investigators to study a wide range of systems; e.g. to identify targets of enhancers harboring GWAS SNPs, targets of developmental stage-specific enhancers, or targets of enhancers that newly arise in specific diseases, such as cancer, diabetes, or neurological disorders. Our new technology is made possible by advances in genome engineering based on the CRISPR/Cas9 system, which enables recruitment of modifying enzymes to specific genomic regions. A notable advantage of this system is the ease of scaling to a comprehensive analysis of a large set of disease-specific enhancers. .